This grant has been focused on understanding the phenomenon of increased genomic instability, manifested as loss-of-heterozygosity (LOH), with advancing replicative age in the budding yeast, S. cerevisiae. In work leading up to the initiation of this R37 grant period, we had discovered that age-associated mitochondrial dysfunction led to the LOH events we observed. We had evidence that this link was the result of reduced production of iron-sulfur clusters (ISCs) by the dysfunctional mitochondria. ISCs are a cofactor for a number of nuclear DNA replication and repair enzymes and we hypothesized that reduction of the ISC cofactor led to impaired DNA repair in old cells. In the past 3.5 years since this grant was awarded, we have made significant progress toward developing a better understanding of early events in the aging process that are upstream of LOH (Hughes and Gottschling, 2012), as well as additional pathways by which age-associated LOH can occur (Lindstrom et al., 2011). Both of these studies were made possible because of the Mother Enrichment Program technology we developed which has allowed us to apply the full potential of tools and breadth of scientific knowledge from S. cerevisiae to study aging (Lindstrom and Gottschling, 2009). While characterizing age-associated phenomena linked to LOH, we made the unexpected and exciting discovery of a new class of proteins (long-lived asymmetrically retained proteins - LARPs) that we believe are highly relevant to understanding the aging process at the cellular level. In fact, one of these LARPs (Pmal) is involved in the earliest steps of the age-associated phenotypes and appears to impact the later age-associated phenotypes of mitochondrial dysfunction (see Progress Report). These findings and the tools we have generated to study them now allow us to explore aspects of aging that have not been tenable before. We believe that study of the LARPs will not only be relevant to yeast replicative aging, but may also affect how we consider the aging process in other cells that undergo repeated asymmetric cell divisions - e.g. stem cells in metazoa. In the next 5 years, I propose to dissect the properties of these unusual proteins and understand what roles they play in the aging process.